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Visualization of aging-associated chromatin alterations with an engineered TALE system

View Article: PubMed Central - PubMed

ABSTRACT

Visualization of specific genomic loci in live cells is a prerequisite for the investigation of dynamic changes in chromatin architecture during diverse biological processes, such as cellular aging. However, current precision genomic imaging methods are hampered by the lack of fluorescent probes with high specificity and signal-to-noise contrast. We find that conventional transcription activator-like effectors (TALEs) tend to form protein aggregates, thereby compromising their performance in imaging applications. Through screening, we found that fusing thioredoxin with TALEs prevented aggregate formation, unlocking the full power of TALE-based genomic imaging. Using thioredoxin-fused TALEs (TTALEs), we achieved high-quality imaging at various genomic loci and observed aging-associated (epi) genomic alterations at telomeres and centromeres in human and mouse premature aging models. Importantly, we identified attrition of ribosomal DNA repeats as a molecular marker for human aging. Our study establishes a simple and robust imaging method for precisely monitoring chromatin dynamics in vitro and in vivo.

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Visualization of aging-associated alterations in genomic repetitive elements by TTALE systems indicating physical attrition of telomeric DNA repeats and NOR-rDNA repeats, as well as decondensation of centromeric DNAs.
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fig9: Visualization of aging-associated alterations in genomic repetitive elements by TTALE systems indicating physical attrition of telomeric DNA repeats and NOR-rDNA repeats, as well as decondensation of centromeric DNAs.

Mentions: Here we report for the first time the direct visualization of heterochromatin disorganization at centromeres during human stem cell aging, further confirming our previous findings7. In addition, we successfully observed telomere shortening3 at the single-cell level both in different human stem cell models of accelerated aging disorders and in vivo in tissues of telomerase-deficient mice. Our most striking finding was the loss of NOR-rDNA TTALE signals in aged human cells. We identified reduction of NOR-rDNA copy number as a novel molecular hallmark of human aging. It should be noted that reduction of rDNA repeats has recently been reported during yeast aging70,71,72. In addition, supporting our conclusion a case report in 1979 suggested aging-associated loss of rDNA in human postmortem myocardium and brain73,74. On the basis of our findings, we propose a model in which human aging is associated with remodeling of genomic repetitive elements, including attrition of telomeric and NOR-rDNA repeats, as well as reorganization of centromeric DNA elements (Figure 9). It should be noted that telomeres, centromeres, and NOR-rDNAs constitute the major components of constitutive heterochromatin within the nucleus75. We also observed a coordinated enhancement of DDRs at telomeric and centromeric loci in human stem cell aging models. Our TTALE-based imaging results provide evidence that human aging involves a complex interplay between genetic and epigenetic instabilities facilitated by changes in 3D chromatin organization.


Visualization of aging-associated chromatin alterations with an engineered TALE system
Visualization of aging-associated alterations in genomic repetitive elements by TTALE systems indicating physical attrition of telomeric DNA repeats and NOR-rDNA repeats, as well as decondensation of centromeric DNAs.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5385610&req=5

fig9: Visualization of aging-associated alterations in genomic repetitive elements by TTALE systems indicating physical attrition of telomeric DNA repeats and NOR-rDNA repeats, as well as decondensation of centromeric DNAs.
Mentions: Here we report for the first time the direct visualization of heterochromatin disorganization at centromeres during human stem cell aging, further confirming our previous findings7. In addition, we successfully observed telomere shortening3 at the single-cell level both in different human stem cell models of accelerated aging disorders and in vivo in tissues of telomerase-deficient mice. Our most striking finding was the loss of NOR-rDNA TTALE signals in aged human cells. We identified reduction of NOR-rDNA copy number as a novel molecular hallmark of human aging. It should be noted that reduction of rDNA repeats has recently been reported during yeast aging70,71,72. In addition, supporting our conclusion a case report in 1979 suggested aging-associated loss of rDNA in human postmortem myocardium and brain73,74. On the basis of our findings, we propose a model in which human aging is associated with remodeling of genomic repetitive elements, including attrition of telomeric and NOR-rDNA repeats, as well as reorganization of centromeric DNA elements (Figure 9). It should be noted that telomeres, centromeres, and NOR-rDNAs constitute the major components of constitutive heterochromatin within the nucleus75. We also observed a coordinated enhancement of DDRs at telomeric and centromeric loci in human stem cell aging models. Our TTALE-based imaging results provide evidence that human aging involves a complex interplay between genetic and epigenetic instabilities facilitated by changes in 3D chromatin organization.

View Article: PubMed Central - PubMed

ABSTRACT

Visualization of specific genomic loci in live cells is a prerequisite for the investigation of dynamic changes in chromatin architecture during diverse biological processes, such as cellular aging. However, current precision genomic imaging methods are hampered by the lack of fluorescent probes with high specificity and signal-to-noise contrast. We find that conventional transcription activator-like effectors (TALEs) tend to form protein aggregates, thereby compromising their performance in imaging applications. Through screening, we found that fusing thioredoxin with TALEs prevented aggregate formation, unlocking the full power of TALE-based genomic imaging. Using thioredoxin-fused TALEs (TTALEs), we achieved high-quality imaging at various genomic loci and observed aging-associated (epi) genomic alterations at telomeres and centromeres in human and mouse premature aging models. Importantly, we identified attrition of ribosomal DNA repeats as a molecular marker for human aging. Our study establishes a simple and robust imaging method for precisely monitoring chromatin dynamics in vitro and in vivo.

No MeSH data available.


Related in: MedlinePlus